hpi2 dylight 650  (Novus Biologicals)

Journal: bioRxiv

Article Title: Glucagon and GLP-1 Accelerate Pseudo-Islet Assembly and Unmask Sex-Specific Islet Fragmentation Dynamics

doi: 10.1101/2025.08.10.669547

Figure Lengend Snippet: a) Workflow: Human donor islets derived from both male and female donors were dissociated into single cells, stained with HIC1-2B4 AF488 (pan-endocrine marker), HIC1-8G12 PE (non-β-endocrine cell marker), and CD9 APC (predominantly δ-cell marker), and sorted into distinct α-, β-, and δ-cell populations via FACS. These populations were then used to form pseudo-islets either from single populations (b) and d) for α-cells ONLY and c) and e) for β-cells ONLY) or combinations thereof (f and g). Pseudo-islets were formed in ultra-low attachment 6-well plates and combined in a 1:10 ratio of endocrine to Mile Sven 1 (MS1) cells (mouse pancreatic endothelial cells). Hormone supplementation consisted of 0.1 µg/mL GCG (pink), 0.1 µg/mL GLP-1 (teal), and 100 nM somatostatin (SST, orange), compared to the control (CTRL, black), with supplements administered every other day. Growth and formation of pseudo-islets depicted in all subpanels are from 2-3 days post-seeding via EPI-light microscopy. Donors were pooled by sex; panels b), c), and f) correspond to female donors and panels d), e), and g) to male donors. Statistical analysis was done by two-way ANOVA with a Benjamini-Hochberg correction for multiple comparisons. P-values are indicated for each comparison; p < 0.05 was considered the cutoff for statistical significance.

Article Snippet: The antibodies used were: HIC1-2B4 A488 conjugated (Novus Biologicals, REF# NBP1-18946AF488) at a 1:100 dilution, HIC1-8G12 PE conjugated (provided by the Grompe lab) at 1:50 dilution, and anti-CD9 APC conjugated to APC (Thermo Fisher Invitrogen, MA1-10307) at a 1:20 dilution.

Techniques: Derivative Assay, Staining, Marker, Control, Light Microscopy, Comparison

Journal: JCI insight

Article Title: HNF1α maintains pancreatic α and β cell functions in primary human islets.

doi: 10.1172/jci.insight.170884

Figure Lengend Snippet: Figure 3. RNA-Seq of HNF1AKD β cells shows that HNF1α regulates insulin secretion, metabolism, developmental pathways, and cell-to-cell signaling in β cells. (A) Schematic of FACS scheme for isolation of transduced live β cells (HPi2+GFP+NTPDase3+) from control and HNF1AKD pseu- doislets for downstream RNA-Seq (n = 4 donors). (B) Fraction of endocrine (HPi2+) cells expressing GFP in sorted samples. (C) HNF1A transcripts per million (TPM) in sequenced samples. (D) Differential expression analysis revealed significantly up- and downregulated genes after HNF1AKD in β cells. Fold change (FC) = 1.5, adjusted P = 0.05. (E) Heatmap of DEGs in β cells after HNF1AKD. (F–H) Significantly downregulated (F) and upregulated (G) Gene Ontology (GO) pathways and downregulated KEGG pathways (H) in HNF1AKD relative to control β cells. *P < 0.05.

Article Snippet: The following primary-secondary conjugated antibodies were used: HPi2-PE/Cy7 (Novus Biologicals NBP1-18946PECY7), NTPDase3-647 (Jean Sévigny’s lab clone hN3-B3S), and CD26-PE (BioLegend 302706) (Supplemental Table 2).

Techniques: RNA Sequencing, Isolation, Control, Expressing, Quantitative Proteomics

Journal: JCI insight

Article Title: HNF1α maintains pancreatic α and β cell functions in primary human islets.

doi: 10.1172/jci.insight.170884

Figure Lengend Snippet: Figure 4. RNA-Seq of HNF1AKD α cells identifies dysregulation of calcium channel complexes and ATPase-coupled transmembrane transport as well as hormone secretion pathways shared with β cells. (A) Schematic of methods for isolation of transduced α cells (HPi2+GFP+CD26+) from control and HNF1AKD pseudoislets for downstream RNA-Seq (n = 4 donors); left image is bright-field, and right image is blue light (488 nm) of human HNF1AKD pseudoislets. Scale bars: 1,000 μm. (B) HNF1A transcripts per million (TPM) in sequenced α cell samples. (C) DEG analysis revealed significantly up- and downregulated genes after HNF1AKD in α cells. FC = 1.5, adjusted P = 0.05. (D) Venn diagram comparing α versus β cell DEGs revealed shared and cell-spe- cific consequences of HNF1AKD. (E and F) Gene Ontology (GO) pathways of shared (E) and α cell (F) enriched DEG sets. (G) Box plots displaying TPM of select DEGs. *P < 0.05.

Article Snippet: The following primary-secondary conjugated antibodies were used: HPi2-PE/Cy7 (Novus Biologicals NBP1-18946PECY7), NTPDase3-647 (Jean Sévigny’s lab clone hN3-B3S), and CD26-PE (BioLegend 302706) (Supplemental Table 2).

Techniques: RNA Sequencing, Isolation, Control